The spatial localization of gene expression can unravel important insights into tissue heterogeneity, functionality and pathological transformations, but the ability to maintain this spatial information remains an enduring challenge in tissue sections routinely used for pathology. Amplification-based spatial gene expression analysis methods provide good sensitivity and specificity but decouple the analyte isolation and biochemical detection steps, making them low throughput and laborious constraints, limiting the translation of the above methods into routine research and clinical practice. Direct probe-based hybridization techniques such as single molecule FISH allow direct visualization of single RNA molecules in their native cellular context but are not amenable on tissue sections in a high throughout manner. In addition, off-target binding of FISH probes and cellular auto-fluorescence can also become a limiting factor in imaging tissue samples. Methods to perform spatially-mapped transcriptome analysis on a tissue section can identify multiple targets simultaneously but they must trade-off between the histologic reference and the quality of recovered biomaterials as staining and manual identification are often needed.
The limitations described above are addressed herein by a special platform to reliably pixelate a tissue section into separate islands of tissue that reside in separate wells and that can be individually analyzed, thereby providing a highly sensitive, reproducible and efficient platform for spatial analysis of tissue. The methods and systems are compatible with on-chip picoliter real-time reverse transcriptase loop mediated isothermal amplification (RT-LAMP) reactions on a histological tissue section, including without any analyte purification, while preserving the native spatial location of the nucleic acid molecules. In an exemplary methodology, the entire process from tissue loading on microchip to results from RT-LAMP, can be carried out in less than two hours. This technique with its ease of use, fast turnaround, and quantitative molecular outputs, is invaluable for a range of applications, including tissue analysis, for researchers and clinicians.